Fig. 2

Neuronal differentiation from control and Alzheimer’s disease induced pluripotent stem cells (AD-iPSCs). a Representative immunofluorescence images show expression of neuronal markers at day 70 of terminal differentiation (TD70): tubulin β 3 class III (TUBB3; green, left panel), microtubule-associated protein 2 (MAP2; red, left panel), TAU (red, middle panel), and neurofilament, heavy polypeptide 200 kDa (NF200; red, right panel). Scale bar = 50 μm. b Differentiation of iPSCs into various neuronal subtypes at TD70 was confirmed by the presence of specific markers: vesicular acetylcholine transporter (VACHT) (cholinergic neurons), glutamic acid decarboxylases 2 and 1 (GAD2/1) (GABAergic neurons), TH (dopaminergic neurons), and vesicular glutamate transporter 1/2 (VGLUT1/2) (glutamatergic neurons). Scale bars = 40 μm and 10 μm as indicated. c Gene expression of neuronal markers from ctr-1, early-onset familial Alzheimer’s disease (fAD-1), and sporadic Alzheimer’s disease (sAD-1) lines at TD70 obtained by qPCR. The expression values were normalized to GAPDH and calculated as a relative amount of messenger RNA versus expression value of neural progenitor cells, which was set to 1. Data are reported as mean ± SEM of three independent measurements. AD-iPSC neurons demonstrate expression pattern similar to that of neuronal cells derived from control iPSCs. d Ultrastructure of a synapse at TD35 with synaptic vesicles (Sv) and synaptic cleft with synaptic junctions (tight junctions, black arrowheads). Docking synaptic vesicles are also observable (white arrowheads). Scale bar = 100 nm. DAPI 4′,6-Diamidino-2-phenylindole